Up-regulation of miR-299 suppressed the invasion and migration of HTR-8/SVneo trophoblast cells partly via targeting HDAC2 in pre-eclampsia.

Pre-eclampsia (PE), a pregnancy-associated disorder, is a major contributor to maternal mortality and morbidity worldwide. Recently, microRNAs (miRNAs) were found to be associated with the pathogenesis of PE. The present study investigated the function of miR-299 in HTR-8/SVneo trophoblast cells and explored its underlying mechanism in the pathogenesis of PE. The miR-299 and histone deacetylase 2 (HDAC2) mRNA expression levels were determined by quantitative real-time PCR. Transwell invasion and wound healing assays were used to measure cell invasive and migratory ability. Luciferase reporter assay was performed to confirm the downstream targets of miR-299. Western blot was performed to measure the protein expression of HDAC2. The expression level of miR-299 in placental tissues from pregnant women with severe PE was significantly higher than that from normal pregnant women. MiR-299 overexpression suppressed the invasion and migration of trophoblast cells; and knock-down of miR-299 promoted the invasion and migration of trophoblast cells. Bioinformatics prediction and luciferase reporter assay confirmed that miR-299 directly targeted the 3' untranslated region of HDAC2. The mRNA expression level of HDAC2 in placental tissues from pregnant women with severe PE was significantly lower than that from normal pregnant women, and was negatively correlated the expression level of miR-299 in placental tissues from pregnant women with severe PE. HDAC2 siRNA transfection suppressed the mRNA and protein expression levels of HDAC in trophoblast cells compared with control group, and HDAC2 siRNA transfection also suppressed the trophoblast cell invasion and migration compared with control group. Enforced expression of HDAC2 in trophoblast cells attenuated the inhibitory effects of miR-299 overexpression on cell invasion and migration. In summary, the results showed the up-regulation of miR-299 in the placental tissues from women with severe PE and miR-299 suppressed the invasion and migration of trophoblast cells partly via targeting HDAC2.

Gao Y ，She R ，Wang Q ，Li Y ，Zhang H ... -  《-》

MiR-221-3p is down-regulated in preeclampsia and affects trophoblast growth, invasion and migration partly via targeting thrombospondin 2.

Preeclampsia (PE) is a common obstetrical disorder characterized by hypertension and proteinuria. The aberrant expression of miR-221-3p in placental tissues has been implicated in the pathogenesis of PE. This study examined the role of miR-221-3p in trophoblast growth, invasion and migration, and explored the underlying mechanisms. Our results showed that miR-221-3p was down-regulated in placental tissues from PE patients compared to heathy controls as measured by quantitative real-time PCR assay. CCK-8 assay, Transwell invasion assay and wound healing assay showed that miR-221-3p overexpression promoted trophoblast (HTR-8/SVneo cells) growth, invasion and migration, and knockdown of miR-221-3p exerted the opposite effects. Flow cytometry experiments further demonstrated that miR-221-3p overexpression inhibited apoptosis, increased cell population at S phase, and decreased cell population at G0/G1 phase of HTR-8/SVneo cells; while miR-221-3p knockdown exerted the opposite effects. Bioinformatics prediction and luciferase reporter assay showed that miR-221-3p targeted the 3' untranslated region of thrombospondin 2 (THBS2), and qRT-PCR and western blot assays revealed that miR-221-3p negatively regulated the expression of THBS2 in HTR-8/SVneo cells. Furthermore, overexpression of THBS2 attenuated the in vitro effects of miR-221-3p overexpression on cell growth, invasion and migration of HTR-8/SVneo cells. The clinical sample analysis showed that the mRNA expression levels of THBS2 was significantly increased in placental tissues from PE patients and was negatively correlated with expression levels of miR-221-3p. In summary, our results demonstrated that miR-221-3p was down-regulated in PE, and the in vitro functional assays revealed that miR-221-3p promoted trophoblast growth, invasion and migration partly via targeting THBS2.

Yang Y ，Li H ，Ma Y ，Zhu X ，Zhang S ，Li J ... -  《-》

Elevated miR-23a impairs trophoblast migration and invasiveness through HDAC2 inhibition and NF-κB activation.

Preeclampsia (PE) is a pregnancy-specific disorder characterized by the onset of hypertension and proteinuria with onset after the 20th week of gestation. The pathogenesis of PE is attributed to increased trophoblast cell death and poor trophoblast migration/invasiveness. This study investigates the function of microRNA-23a (miR-23a) in PE and its effects on migration and invasion of trophoblast cells HTR-8/SVneo. We found higher expression of miR-23a in placental tissue samples from PE pregnant women compared to samples from normal pregnant women. Enhancing miR-23a expression by its specific mimic reduced HTR-8/SVneo cell migration and invasion and increased HTR-8/SVneo cell apoptosis. The dual-luciferase reporter gene assay revealed miR-23a binding with HDAC2. We found that HDAC2 was poorly expressed in placental tissue samples from PE pregnant women, and its expression correlated inversely with miR-23a expression. HTR-8/SVneo cells showed diminished HDAC2 expression upon miR-23a elevation and increased HDAC2 expression upon miR-23a inhibition. Lentivirus-mediated HDAC2 knockdown mimicked the effects of miR-23a on HTR-8/SVneo cells and led to NF-κB activation. Similarly, HDAC2 overexpression and NF-κB inhibition both abrogated the effects of miR-23a on HTR-8/SVneo cells, suggesting that miR-23a reduced HTR-8/SVneo cell migration and invasion and increased HTR-8/SVneo cell apoptosis by HDAC2 inhibition and NF-κB activation. In summary, these results support a novel role of miR-23b in invasion and apoptosis of trophoblast cells, and imply that targeting miR-23b may be a new avenue for treating PE.

Fan Y ，Dong Z ，Zhou G ，Fu J ，Zhan L ，Gao M ，Zhu L ，Zhang Y ... -  《-》

Upregulated long noncoding RNA Linc00261 in pre-eclampsia and its effect on trophoblast invasion and migration via regulating miR-558/TIMP4 signaling pathway.

Pre-eclampsia (PE) is a leading cause of maternal and perinatal morbidity and mortality but the exact underlying mechanisms of PE pathogenesis remain elusive. Accumulated data suggested that the long noncoding RNAs (lncRNAs) play important roles in the pathogenesis of PE. The present study identified the changes of lncRNA Linc00261 in PE and its effects on trophoblasts invasion and migration. Our results showed that the expression of Linc00261 was upregulated in placental tissues of PE women compared with those of healthy pregnant women. Overexpression of Linc00261 suppressed cell invasion and migration, induced cell apoptosis, and caused cell-cycle arrest at G0 /G1 phase of HTR-8/SVneo cells; while knockdown of Linc00261 had the opposite effects on the HTR-8/SVneo cells. Mechanistic studies showed Linc00261 functioned as a competing endogenous RNA for miR-558 in HTR-8/SVneo cells, and miR-558 was negatively regulated by Linc00261. The expression level of miR-558 in the PE group was significantly lower than the control group, and the expression level of Linc00261 was negatively correlated with the expression level of miR-558 in the placental tissues of women with PE. Furthermore, miR-558 was found to negatively regulate the expression of TIMP metallopeptidase inhibitor 4 (TIMP4) via targeting the 3' untranslated region in the HTR-8/SVneo cells. Overexpression of miR-558 increased HTR-8/SVneo cell invasion and migration, which was attenuated by TIMP4 overexpression. More importantly, both overexpression of miR-558 and knockdown of TIMP4 partially reversed the suppressive effects of Linc00261 overexpression on cell invasion and migration of HTR-8/SVneo cells. Collectively, our results for the first time showed the upregulation of Linc00261 in the placental tissues of severe PE patients. The mechanistic results indicated that Linc00261 exerted the suppressive effects on the trophoblast invasion and migration via targeting miR-558/TIMP4 axis, which may involve in the pathogenesis of PE.

Cheng D ，Jiang S ，Chen J ，Li J ，Ao L ，Zhang Y ... -  《-》

The decreased lncRNA ZEB2-AS1 in pre-eclampsia controls the trophoblastic cell line HTR-8/SVneo's invasive and migratory abilities via the miR-149/PGF axis.

Pre-eclampsia (PE) is a pregnancy disease that causes maternal death and threatens the health of newborns. Accumulating evidence has revealed the essential roles of long noncoding RNAs (lncRNAs) in the progression of PE. The present investigation determined lncRNA ZEB2 antisense RNA 1 (ZEB2-AS1) expression in PE and looked into the potential role of ZEB2-AS1 in modulating trophoblastic cell functions. Quantitative real-time polymerase chain reaction evaluated gene expression. Western blot analyzed the placental growth factor (PGF) protein level. Cell counting kit-8 and Transwell invasion assays assessed the proliferative and invasive abilities of placental trophoblast cells, respectively. Wound healing assay determined cell migratory potentials. Dual-luciferase reporter assay assessed the targeting relationship among ZEB2-AS1, miR-149, and PGF. Downregulation of lncRNA ZEB2-AS1 was detected in placentas from patients with PE when compared with those from normal pregnancies. Moreover, ZEB2-AS1 upregulation markedly promoted proliferative, migratory, and invasive potentials in HTR-8/SVneo cells, while knockdown of ZEB2-AS1 had the opposite effects. The effects on HTR-8/SVneo cells mediated by ZEB2-AS1 was correlated with the miR-149/PGF axis. These findings indicate that ZEB2-AS1 contributes to PE progression by affecting cell proliferative and invasive capacities via the miR-149/PGF axis in HTR-8/SVneo cells. In sum, we identified that ZEB2-AS1 was a novel aberrantly expressed lncRNA in the placentas of PE patients and lncRNA ZEB2-AS1 modulated trophoblastic cell line HTR-8/SVneo's proliferative and invasive potentials via targeting the miR-149/PGF axis.

Gao Y ，Guo X ，Li Y ，Sha W ，She R ... -  《-》